Nontoxic clostridium perfringens and clostridium septicum fusion protein vaccine and production method thereof

A technology of Clostridium perfringens and fusion protein, applied in vaccines, multivalent vaccines, veterinary vaccines, etc., can solve problems such as incomplete antigenicity of antigenic proteins, biosafety risks, and impact on immunogenicity, so as to avoid Effects of antigenic protein immunogenicity, maintenance of immunogenicity, and effect of reducing biosafety risks

CN110051834APending Publication Date: 2019-07-26CHINA INST OF VETERINARY DRUG CONTROL
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Publication Date
2019-07-26

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Abstract

The invention relates to a nontoxic clostridium perfringens and clostridium septicum fusion protein vaccine and a production method thereof. The fusion protein vaccine prepared by the method adopts aclostridium perfringens epsilon toxin which is subjected to codon optimization and contains three amino acid mutations, and spoilage which contains four amino acid mutations and eleven amino acid deletions, namely not only kept the integrity and spatial conformation of the natural toxin are to the maximum extent, but also the biological safety hazard caused by single amino acid mutation is avoided. The vaccine also has the advantages of simple preparation process, excellent vaccine efficacy and the like, greatly reduces the biological safety risk in the vaccine production process. Compared with the current commercialized sheep fast-epidemic and sheep enterotoxemia inactivated vaccines in China, the vaccine is an ideal candidate vaccine for upgrading existing clostridium perfringens and clostridium septicum vaccines in China; in addition, when combined vaccines are prepared by the vaccine together with other antigens, the combined vaccine can be prepared without increasing the use doseof the combined vaccine.
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Description

technical field

[0001] The invention relates to an avirulent Clostridium perfringens and Clostridium putrefaction fusion protein vaccine and a production method thereof. It belongs to the field of veterinary biological products. Background technique

[0002] Clostridium perfringens and Clostridium putrefaction are anaerobic bacteria that can cause disease in humans and various animals, and are extremely harmful to human health and livestock and poultry breeding. Among them, Clostridium perfringens is one of the main pathogens of traumatic gas gangrene and human food poisoning, sudden gangrene of sheep, lamb dysentery, necrotic enteritis of cattle and sheep, and enterotoxemia of cattle and sheep. The main pathogen of the epidemic. The pathogenic factors of both Clostridia are exotoxins secreted by the bacteria. Clostridium perfringens can be divided into A, B, C, D, E according to the types of four main lethal exotoxins α (CPA), β (CPB), ε (ETX) and ι (CPI) produced. Five...

Examples

Embodiment 1

[0058] ——Construction and identification of Escherichia coli BL / EC strain

[0059] 1. Gene synthesis

[0060] The application is based on the natural gene sequences of Clostridium perfringens ETX and Clostridium putrefaction CSA, after codon optimization, the ETX coding gene containing 3 amino acid mutations (Y30A, H106P and Y196A), the ETX coding gene containing 4 amino acid mutations ( C54L, N264A, H269A and W310A) and 11 amino acid deleted CSA coding genes were tandem. At the same time, the gene encoding the 6×His tag was added to the C-terminus of the fusion protein. The gene sequence GETX was synthesized by chemical synthesis m3 CSA M4△11 , containing a total of 2166 nucleotides, the corresponding protein is called rETX m3 CSA M4△11 . The specific nucleic acid sequence is shown in SEQ ID No.1, and the amino acid sequence is shown in SEQ ID No.2.

[0061] Sequence 1:

[0062]

[0063]

[0064]

[0065] Sequence 2:

[0066]

[0067]

[0068] 2. Expre...